PROJECT SUMMARY Malignant melanoma is an aggressive disease for which there are limited therapeutic options. In particular, individuals over the age of 55 have a much poorer prognosis for melanomas of equal grade and stage, compared to younger individuals. To date, no one has addressed the reasons for this disparity in melanoma aggressiveness and metastasis in aging individuals. Our long-term objective is test the hypothesis that the aging microenvironment drives the local aggressiveness and systemic dissemination of melanoma cells. In previous work we identified a critical signaling pathway that modulates melanoma invasiveness, by the secreted factor Wnt5a. We have found that melanomas with high Wnt5A have a mesenchymal phenotype, an increased propensity to migrate and invade, and are significantly associated with poorer prognosis. In preliminary data for this application we show that co-culture of melanomas with fibroblasts from aged individuals, but not young individuals, induces them to express high Wnt5a levels, a mesenchymal phenotype, and increased invasiveness. Interestingly, we have also discovered that melanomas co-cultured with aged but not young fibroblasts show evidence for a senescent-like phenotype. Our preliminary data implicate the aging-regulated hormone Klotho in this switch to an invasive, senescent- like phenotype induced by aged fibroblasts. We find that aged fibroblasts consistently decrease their expression of Klotho, and that Klotho is a potent inhibitor of Wnt5A signaling and senescence. We hypothesize that the decreased expression of Klotho within the aged tumor micro-environment contributes to increased Wnt5A signaling and increased invasiveness in melanomas in the elderly. The proposed studies will lead to an improved understanding of the role of Klotho and the aging micro-environment in the increased aggressiveness and invasiveness seen in melanomas in patients over the age of 55. It is expected that elucidating the role of Klotho and Wnt5a in melanomas in aging will lead to improved therapy.